DESCRIPTION:(PROVIDED BY APPLICANT) We have recently identified cyclin G as being overexpressed in breast cancer cells using differential screening. We found that cyclin G overexpression was a frequent occurrence in human breast cancer specimens. Cyclin G, a recent addition to the cyclin family, was identified as a transcriptional target of the tumor suppressor, p53. Although the biological role(s) of cyclin G has not yet been defined, transactivation of cyclin G by p53 is intriguing since other cyclins have been shown to play crucial roles in the control of cell cycle and growth. Transcriptional induction of cyclin G by p53 was only observed in normal or non-tumorigenic cells. Preliminary data suggest that following DNA damage in normal mammary epithelial cells, as well as in fibroblasts, cyclin G is triggered to cluster in discrete nuclear DNA replication foci that contain replication-associated proteins such as PCNA. Initial characterization of cyclin G suggests that there is a link between p53-mediated tumor suppression and DNA replication. Additionally, we observed that retrovirus-mediated overexpression of cyclin G induced cell cycle arrest in 10.1 fibroblasts, which contain no p53, suggesting that overexpression of cyclin G in p53-null cells mimics the tumor suppressive role of p53. The overall goal of this revised project is to characterize the phenotypic changes and the undefined role(s) of cyclin G related to mammary cell proliferation in vitro and in vivo. We will also analyze the molecular basis of increased transcription of cyclin G in breast cancer cells by further localizing the responsive region of the cyclin G promoter. In addition, we will determine whether the enhanced protein stability of cyclin G can contribute to cyclin G overexpression in breast cancer cells by using a chimeric molecule with cyclin G fused to luciferase protein. A variety of p53 mutants will be analyzed in an effort to identify and define p53 determinants required for cyclin G induction. Lastly, we will generate transgenic mice, expressing cyclin G, and cross these mice with other existing cyclin transgenic mice including cyclin D, p2 1-/-, and ErbB2-overexpressing mice as well as p53 null mice to identify the role of cyclin G in vivo. Better understanding of the roles of cyclin G in cell growth control should offer a unique opportunity to study a novel mechanism for p53-mediated tumor suppression and also develop a novel approach targeting for mammary cell transformation.